PROJECT SUMMARY Type 1 diabetes (T1D) is an autoimmune disease that affects millions of people worldwide. The incidence of T1D is rising, especially in young children. Although significant progress has been made to predict who is at risk for developing T1D, there are no effective therapies to prevent this disease. Both genetic and environmental factors contribute to the risk of developing T1D. Certain human leukocyte antigen (HLA) haplotypes dominantly protect against the development of T1D, yet the mechanism of this remarkable protection from autoimmunity is not well-understood. NOD mice, the most widely used model of T1D, do not express a major histocompatibility complex (MHC) class II E molecule. Transgenic expression of the MHCII E molecule in NOD mice (E?16/NOD) completely prevents T1D, mirroring dominant HLA protection from T1D in humans. Using these E?16/NOD mice as a model of dominant genetic protection from T1D, we recently demonstrated that protection from autoimmunity operates by the immune system shaping the early-life commensal microbiota. Modeling of HLA class II dominant protection from T1D in murine models may provide critical insights to support our long-term goal of developing microbiota-based therapies to prevent T1D in humans. Due to the complexity and high levels of variability of the intestinal microbiome, determining the specific microbial strains that are modulated by the immune system is problematic. The development of gnotobiotic mice with defined adult microbial communities has been an important advance in the field because they simplify the complexity and variability of the system and allow for well-controlled, mechanistic studies. However, a gnotobiotic mouse model to study pediatric disease is lacking. We have leveraged the E?16/NOD mouse model of genetic protection from T1D to generate a new gnotobiotic mouse model of the early-life microbiome which we call Pediatric Community or ?PedsCom-A?. PedsCom-A is a consortium of 9 bacterial strains isolated from the intestine of pre-weaning diabetes-protected E?16/NOD mice. We hypothesize that MHCII molecules play a major role in shaping the intestinal microbiome early in development, and these microbes in turn impact the development of the immune system to prevent T1D. Aim 1 examines the mechanisms of interaction and colonization dynamics of these 9 bacteria in PedsCom-A colonized NOD mice and NOD mice expressing the MHCII E molecule (E?16/NOD). Aim 2 examines whether PedsCom-A microbes prevents T1D in NOD and E?16/NOD mice. Successful completion of these aims will provide critical information on which early-life microbes are influenced by expression of the MHCII E molecule to generate a diabetes-protective microbiome, and whether these 9 microbes that constitute PedsCom are sufficient to prevents T1D in diabetes-prone NOD mice. In addition, PedsCom-A mice are an innovative tool for investigating early-life host-microbiota interactions.